In phase I, Nanosphere has demonstrated feasibility for a gold nanoparticle-based detection system for biological threat agents using a Bacillus subtilis model system. In phase II, the detection sensitivity of this platform will be increased by applying nanoparticle-based bio-barcode technology and automated microfluidic assay processing. The goal is to achieve detection of 60 target copies in < 2 hours total assay time without using enzyme-based target (e.g. PCR) or signal amplification. This new array based assay and detection system will provide clinical diagnostics labs a lower cost alternative to current real time PCR detection systems, and additionally, multiplex detection capabilities for infectious agent identification. In year 2, we will develop and test the sensitivity and specificity of assays for biological threat agents (e.g. Bacillus anthracis, Yersinia Pestis) and extend the assay to the detection of methicillin resistant Staphylococcus aureus. Given the urgent need for rapid and inexpensive tests for biological threat agent identification at the point-of-site, Nanosphere also will integrate bio-barcode technology with a novel simple and rapid high sensitivity colorimetric assay for nucleic acid detection developed at Nanosphere. This platform will utilize simplified assay procedures along with a visual colorimetric readout to achieve detection of 100 target copies in < 30 minutes for point-of-site detection applications, and especially, biological threat agent detection. In year 2, we will develop and test the sensitivity and specificity of the biological threat agents Bacillus anthracis and Yersinia pestis. [unreadable] [unreadable]